Project Summary/Abstract: Recent studies of addiction have highlighted several regions of the brain that are thought to be involved in goal directed and drug seeking behaviors. The specific neuronal classes involved in the regulation of these behaviors are beginning to be identified, and attempts to profile the molecular changes in these cell types occurring as a consequence of addiction have met with some success. While these studies demonstrate that it is possible to discover changes in cell types that are correlated with, and in some cases contribute to, addiction, our knowledge in this area is fragmentary and incomplete. This is due to technical obstacles that have faced this field for decades, and that have recently been overcome by novel methodologies developed in our laboratories. The objective of this program of research is to identify all of the changes in gene expression and accompanying alterations in epigenetic regulation that contribute to the addictive state in cell types known to be important in the neural circuitry controlling addiction. The approach we will take in this program is to: 1) employ TRAP methodology and bacTRAP transgenic mouse lines to comprehensively profile the translated mRNAs from fifteen mouse CNS cell types that are components of the neural circuitry controlling addiction;2) to collect translational profiles from each of these cell types in mice exposed to cocaine, methylphenidate, and methamphetamine 3) perform in depth comparative analysis of these resulting microarray datasets to identify changes in gene expression that accompany the addictive state in each cell type;4) to concurrently isolate nuclei from each cell type and map cell specific sites of mC and hmC modification to the neuronal genomes;5) to conduct follow up studies on genomic loci identified in the preceding aims to map additional epigenetic regulatory events by ChIP assays using H3K9m2 and H3K27m3 specific antibodies to identify genomic loci associated with suppression of gene expression in euchromatin. This project will provide information and experimental animals that will stimulate addiction research in a broad spectrum of laboratories, and provide materials and a paradigm for comprehensive studies of these same neural circuits under other experimental conditions. As such, this program will have an enormous impact on modern molecular neurobiology, and on the development of novel targets for pharmacological interventions in CNS disorders. PUBLIC HEALTH RELEVANCE: According to the National Institute on Drug Abuse, "Estimates of the total overall costs of substance abuse in the United States ..... exceed half a trillion dollars annually". The goals of the project are to comprehensively profile molecular changes occurring in specific cell types in regions associated with addiction, and to concurrently map epigenetic changes occurring in these cell types. It will stimulate in depth research into cell specific molecular events that are responsible for establishment of the addictive state, and identify novel targets for the development of new therapies for drug addiction.